Structure-based design of covalent nanobody binders for a thermostable green fluorescence protein
The use of green fluorescence protein (GFP) has advanced numerous areas of life sciences. An ultra-thermostable GFP (TGP), engineered from a coral GFP, offers potential advantages over traditional jellyfish-derived GFP because of its high stability. However, owing to its later discovery, TGP lacks t...
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| Format: | Article |
| Language: | English |
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China Science Publishing & Media Ltd.
2024-12-01
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| Series: | Acta Biochimica et Biophysica Sinica |
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| Online Access: | https://www.sciengine.com/doi/10.3724/abbs.2024233 |
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| author | Yue Zhihao Li Yanfang Cai Hongmin Yao Hebang Li Dianfan Ni Aimin Li Tingting |
| author_facet | Yue Zhihao Li Yanfang Cai Hongmin Yao Hebang Li Dianfan Ni Aimin Li Tingting |
| author_sort | Yue Zhihao |
| collection | DOAJ |
| description | The use of green fluorescence protein (GFP) has advanced numerous areas of life sciences. An ultra-thermostable GFP (TGP), engineered from a coral GFP, offers potential advantages over traditional jellyfish-derived GFP because of its high stability. However, owing to its later discovery, TGP lacks the extensive toolsets available for GFP, such as heavy chain-only antibody binders known as nanobodies. In this study, we report the crystal structure of TGP in complex with Sb92, a synthetic nanobody identified from a previous in vitro screening, revealing Sb92’s precise three-dimensional epitope. This structural insight, alongside the previously characterized Sb44-TGP complex, allows us to rationally design disulfide bonds between the antigen and the antibody for tighter interactions. Using biochemical analysis, we identify two bridged complexes (TGP A18C-Sb44 V100C and TGP E118C-Sb92 S57C), with the TGP-Sb92 disulfide pair showing high resistance to reducing agents. Our study expands the toolkit available for TGP and should encourage its wider applications. |
| format | Article |
| id | doaj-art-e7c7cb2fdf354b5896ca6d3e93c78f81 |
| institution | Kabale University |
| issn | 1672-9145 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | China Science Publishing & Media Ltd. |
| record_format | Article |
| series | Acta Biochimica et Biophysica Sinica |
| spelling | doaj-art-e7c7cb2fdf354b5896ca6d3e93c78f812025-08-20T03:47:12ZengChina Science Publishing & Media Ltd.Acta Biochimica et Biophysica Sinica1672-91452024-12-01571363137010.3724/abbs.202423320d259ccStructure-based design of covalent nanobody binders for a thermostable green fluorescence proteinYue Zhihao0Li Yanfang1Cai Hongmin2Yao Hebang3Li Dianfan4Ni Aimin5Li Tingting6["Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China","University of Chinese Academy of Sciences, Beijing 100049, China"]["Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China"]["Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China","State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201210, China"]["Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China","School of Life Science and Technology, ShanghaiTech University, Shanghai 201210, China"]["Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China"]["Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China"]["Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China"]The use of green fluorescence protein (GFP) has advanced numerous areas of life sciences. An ultra-thermostable GFP (TGP), engineered from a coral GFP, offers potential advantages over traditional jellyfish-derived GFP because of its high stability. However, owing to its later discovery, TGP lacks the extensive toolsets available for GFP, such as heavy chain-only antibody binders known as nanobodies. In this study, we report the crystal structure of TGP in complex with Sb92, a synthetic nanobody identified from a previous in vitro screening, revealing Sb92’s precise three-dimensional epitope. This structural insight, alongside the previously characterized Sb44-TGP complex, allows us to rationally design disulfide bonds between the antigen and the antibody for tighter interactions. Using biochemical analysis, we identify two bridged complexes (TGP A18C-Sb44 V100C and TGP E118C-Sb92 S57C), with the TGP-Sb92 disulfide pair showing high resistance to reducing agents. Our study expands the toolkit available for TGP and should encourage its wider applications.https://www.sciengine.com/doi/10.3724/abbs.2024233disulfide engineeringrational designsynthetic nanobodythermostable green fluorescence protein |
| spellingShingle | Yue Zhihao Li Yanfang Cai Hongmin Yao Hebang Li Dianfan Ni Aimin Li Tingting Structure-based design of covalent nanobody binders for a thermostable green fluorescence protein Acta Biochimica et Biophysica Sinica disulfide engineering rational design synthetic nanobody thermostable green fluorescence protein |
| title | Structure-based design of covalent nanobody binders for a thermostable green fluorescence protein |
| title_full | Structure-based design of covalent nanobody binders for a thermostable green fluorescence protein |
| title_fullStr | Structure-based design of covalent nanobody binders for a thermostable green fluorescence protein |
| title_full_unstemmed | Structure-based design of covalent nanobody binders for a thermostable green fluorescence protein |
| title_short | Structure-based design of covalent nanobody binders for a thermostable green fluorescence protein |
| title_sort | structure based design of covalent nanobody binders for a thermostable green fluorescence protein |
| topic | disulfide engineering rational design synthetic nanobody thermostable green fluorescence protein |
| url | https://www.sciengine.com/doi/10.3724/abbs.2024233 |
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